Insecticide résistance study and biological responses to climate changesof Aedes aegypti mosquito, the dengue, chikungunya and zika vector in Guadeloupe

La Guadeloupe fait partie des pays où la Dengue est endémique avec des épidémies tous les 2 à 3 ans. Depuis 3 ans, d'autres arboviroses sont apparues sur le continent américain avec le Chikungunya en 2013 puis le Zika en 2015, causant d'importantes épidémies notamment en Guadeloupe. Le seul vecteur reconnu de ces maladies en Guadeloupe est le moustique Aedes aegypti. Il n'y a pas de vaccin, ni de traitements spécifiques contre ces infections et les moyens de prévention contre ces maladies passent par la surveillance et le contrôle des populations de moustiques sur le terrain. Les méthodes de surveillance sont basées le plus souvent sur l'analyse d'indices larvaires, parfois controversés. De plus, les moyens de contrôle des vecteurs ont longtemps été basés sur l'utilisation massive d'insecticides chimiques entraînant la résistance des moustiques à ces produits. Ce travail de thèse s'est donc articulé autour de deux grands axes devant permettre d'améliorer la prévention et le contrôle de ces arboviroses: i) la recherche d'un nouvel outil de surveillance des populations vectrices, basé sur la physiologie des femelles adultes et ii) l'évaluation des niveaux et l'étude de certains mécanismes de résistance à trois insecticides chimiques, le Téméphos, le Malathion (utilisés dans le passé) et la Deltaméthrine (utilisée actuellement). Un modèle de surveillance des populations vectrices basé sur les taux de parité en lien avec l'espérance de vie des femelles, en fonction des températures a été développé, et des pistes sur les situations entomologiques les plus à risques se sont dessinées. Les épreuves de résistance effectuées sur des larves de moustiques de Guadeloupe ont globalement révélé de forts niveaux de résistance au Téméphos et de faibles niveaux de résistance au Malathion. Les tests adulticides ont mis en évidence une résistance modérée des femelles à la Deltaméthrine. Les investigations moléculaires ont démontré des fréquences alléliques très élevées pour les mutations Kdr V1016I et F1534C connues pour être liées à la résistance aux pyréthrinoïdes. De plus, l'évaluation des niveaux d'expression constitutifs de certains gènes de détoxification a révélé des surexpressions significatives des populations testées par rapport à la souche sensible Bora-Bora, pour la carboxy-choline-estérase CCEAE3A, quatre cytochromes P450 à mono-oxygénases (014614, CYP6M11, CYP6BB2 et CYP9J23) et la glutathione-S-transférase GSTE2.Guadeloupe is an endemic country for Dengue with epidemics every 2 to 3 years. In the past 3 years, other arboviruses have reached the Americas with Chikungunya virus in 2013 and Zika virus in 2015, causing major epidemics including in Guadeloupe. The only known vector of these diseases in Guadeloupe is the mosquito Aedes aegypti. As there is no vaccine nor specific treatment against these infections, prevention against these diseases is achieved through the monitoring and control of mosquito populations. Monitoring methods are based mostly on larval indices, with sometimes controversial results. In addition, vector control methods are based since a very long time on the massive use of chemical insecticides, causing mosquito resistance to these products. This work has therefore focused on two main areas to improve the prevention and control of these arboviruses: i) the search of a new vector population monitoring tool, based on the physiology of adult females and ii) the assessment of the resistance levels and mechanisms regarding three chemical insecticides, Temephos, Malathion (used in the past) and Deltamethrin (currently used). A vector population monitoring model based on females life expectancy as a function of parity rates and according to temperatures has being developed, and tracks on the entomological situations most at risk have emerged. Insecticide resistance tests performed on mosquito larvae have generally found strong Temephos resistance levels and low resistance to Malathion. Adulticide tests showed a moderate resistance of females to Deltamethrin. Molecular investigations have shown very high allelic frequencies for kdr mutations V1016I and F1534C, known to be associated with pyrethroid resistance. Moreover, the evaluation of constitutive expression levels of some detoxification genes revealed significant overexpression in tested Aedes aegypti populations compared to the susceptible Bora-Bora strain, for the carboxy-choline-esterase CCEAE3A, four cytochrome P450 mono-oxygenases (014614, CYP6M11, CYP6BB2 and CYP9J23) and the glutathione-S-transferase GSTE2

At the Origin of a Worldwide Invasion: Unraveling the Genetic Makeup of the Caribbean Bridgehead Populations of the Dengue Vector Aedes aegypti

International audienceHuman-driven global environmental changes have considerably increased the risk of biological invasions, especially the spread of human parasites and their vectors. Among exotic species that have major impacts on public health, the dengue fever mosquito Aedes aegypti originating from Africa has spread worldwide during the last three centuries. Although considerable progress has been recently made in understanding the history of this invasion, the respective roles of human and abiotic factors in shaping patterns of genetic diversity remain largely unexplored. Using a genome-wide sample of genetic variants (3,530 ddRAD SNPs), we analyzed the genetic structure of Ae. aegypti populations in the Caribbean, the first introduced territories in the Americas. Fourteen populations were sampled in Guyane and in four islands of the Antilles that differ in climatic conditions, intensity of urbanization, and vector control history. The genetic diversity in the Caribbean was low (He = 0.14-0.17), as compared with a single African collection from Benin (He = 0.26) and site-frequency spectrum analysis detected an ancient bottleneck dating back ∼300 years ago, supporting a founder event during the introduction of Ae. aegypti. Evidence for a more recent bottleneck may be related to the eradication program undertaken on the American continent in the 1950s. Among 12 loci detected as FST-outliers, two were located in candidate genes for insecticide resistance (cytochrome P450 and voltage-gated sodium channel). Genome-environment association tests identified additional loci associated with human density and/or deltamethrin resistance. Our results highlight the high impact of human pressures on the demographic history and genetic variation of Ae. aegypti Caribbean populations

Multiple insecticide resistance in Culex quinquefasciatus populations from Guadeloupe (French West Indies) and associated mechanisms.

West Nile (WN) virus has been detected in Guadeloupe since 2002. Even if no WN human cases have been detected so far, mosquitoes from Culex genus especially Culex quinquefasciatus are recognized as potential WN vectors in Guadeloupe. To evaluate the impact of local vector control activities on this mosquito species we assessed the resistance levels of Cx. quinquefasciatus populations from three different sites from Guadeloupe (Abymes, Saint François and Gourbeyre) to malathion, temephos and deltamethrin. In addition, the frequencies of the L1014F kdr and the G119S ace-1 mutations were established in Cx. quinquefasciatus populations, as well as the constitutive expressions of five cytochrome P450 genes. Mosquito populations tested displayed high resistance to deltamethrin, moderate resistance to malathion (Abymes, Gourbeyre) and low resistance to temephos (Abymes et Gourbeyre). Molecular analyses revealed high frequencies of both L1014F kdr and G119S ace-1 mutations in Cx. quinquefasciatus populations, as well as overexpression of cytochrome P450 genes CYP9J45, CYP9J40 and CYP6AA7. Finally, deltamethrin resistance and knock-down rates were strongly correlated with the frequency of the resistant kdr and ace-1 alleles, as well as with CYP9J40 overexpression. These results should be taken into account to refine the current vector control strategies to prevent the appearance of Cx. quinquefasciatus-borne diseases in Guadeloupe

Survival of females of <i>Aedes aegypti</i> in laboratory conditions at constant temperatures of 24°C, 27°C and 30°C plotted against the age of females.

<p>Survival of females of <i>Aedes aegypti</i> in laboratory conditions at constant temperatures of 24°C, 27°C and 30°C plotted against the age of females.</p

Graphical representation of the biological events occurring in the <i>Aedes aegypti</i> female life-cycle at optimal (A) and lower (B) temperatures in relation with the dengue and chikungunya virus infections, amplification and transmission.

<p>GC = Gonotrophic Cycle, EIP = Extrinsic Incubation period.</p

Linear regression representing the mean durations of the Gonotrophic Cycles as a function of temperatures (<i>y</i> = −0.4998<i>x</i> + 20.368, r² = 0.98).

<p>Linear regression representing the mean durations of the Gonotrophic Cycles as a function of temperatures (<i>y</i> = −0.4998<i>x</i> + 20.368, r² = 0.98).</p

Improvement of mosquito identification by MALDI-TOF MS biotyping using protein signatures from two body parts

Abstract Background Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry technology (MALDI-TOF MS) is an innovative tool that has been shown to be effective for the identification of numerous arthropod groups including mosquitoes. A critical step in the implementation of MALDI-TOF MS identification is the creation of spectra databases (DB) for the species of interest. Mosquito legs were the body part most frequently used to create identification DB. However, legs are one of the most fragile mosquito compartments, which can put identification at risk. Here, we assessed whether mosquito thoraxes could also be used as a relevant body part for mosquito species identification using a MALDI-TOF MS biotyping strategy; we propose a double DB query strategy to reinforce identification success. Methods Thoraxes and legs from 91 mosquito specimens belonging to seven mosquito species collected in six localities from Guadeloupe, and two laboratory strains, Aedes aegypti BORA and Aedes albopictus Marseille, were dissected and analyzed by MALDI-TOF MS. Molecular identification using cox1 gene sequencing was also conducted on representative specimens to confirm their identification. Results MS profiles obtained with both thoraxes and legs were highly compartment-specific, species-specific and species-reproducible, allowing high identification scores (log-score values, LSVs) when queried against the in-house MS reference spectra DB (thorax LSVs range: 2.260–2.783, leg LSVs range: 2.132–2.753). Conclusions Both thoraxes and legs could be used for a double DB query in order to reinforce the success and accuracy of MALDI-TOF MS identification

Expectations of life (in days) and expectations of infective life (in days) for <i>An</i>. <i>gambiae</i> and for <i>Ae</i>. <i>aegypti</i> from the parity rates.

<p><i>An</i>. <i>gambiae</i> models are based on Coz & al. (1961) and Garret-Jones and Grab (1964) publications.</p><p>^aFor <i>Ae</i>. <i>aegypti</i> and dengue virus EIP is estimated from Carrington et al. (2013)</p><p>Expectations of life (in days) and expectations of infective life (in days) for <i>An</i>. <i>gambiae</i> and for <i>Ae</i>. <i>aegypti</i> from the parity rates.</p

Tukey-type test results for comparison among proportion of blood feeding females, parity rate of blood-engorged females, overall parity rates, survival at 25 days of age, proportion of females having 2 GCs and the proportion of females having 3 GCs, at the different experimental temperatures.

<p>* for significant differences</p><p>Tukey-type test results for comparison among proportion of blood feeding females, parity rate of blood-engorged females, overall parity rates, survival at 25 days of age, proportion of females having 2 GCs and the proportion of females having 3 GCs, at the different experimental temperatures.</p

Expectation of infective life (I_i) in days for 50% of the females of <i>Aedes aegypti</i> females according to the EIP at constant temperatures of 24°C, 27°C and 30°C, plotted against the parity rates.

<p>Expectation of infective life (I_i) in days for 50% of the females of <i>Aedes aegypti</i> females according to the EIP at constant temperatures of 24°C, 27°C and 30°C, plotted against the parity rates.</p